Genetic Diversity and Detection of Atypical Porcine Pestivirus Infections

2021 ◽  
Author(s):  
Kylee M Sutton ◽  
Christian W Eaton ◽  
Tudor Borza ◽  
Thomas E Burkey ◽  
Benny E Mote ◽  
...  
Keyword(s):  
Rna Virus ◽  
Protein Translation ◽  
Template Switching ◽  
Substantial Variation ◽  
Functional Importance ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Viral Cdna ◽  
Viral Polyprotein ◽  
Congenital Tremor

Abstract Atypical porcine pestivirus (APPV), an RNA virus member of the Flaviviridae family, has been associated with congenital tremor in newborn piglets. Previously reported qPCR-based assays were unable to detect APPV in novel cases of congenital tremor originated from multiple farms from U.S. Midwest (MW). These assays targeted the viral polyprotein coding genes, which were shown to display substantial variation, with sequence identity ranging from 58.2 to 70.7% among 15 global APPV strains. In contrast, the 5’ UTR was found to have a much higher degree of sequence conservation. In order to obtain the complete 5’ UTR of the APPV strains originated from MW, the 5’ end of the viral cDNA was obtained by using template switching approach followed by amplification and dideoxy sequencing. Eighty one percent of the 5’UTR was identical across 14 global and 5 MW strains with complete, or relatively complete 5’ UTR. Notably, some of the most highly conserved 5’UTR segments overlapped with potentially important regions of an internal ribosome entry site (IRES), suggesting their functional role in viral protein translation. A newly designed single qPCR assay, targeting 100% conserved 5’UTR regions across 19 strains, was able to detect APPV in samples of well documented cases of congenital tremor which originated from five MW farm sites (1-18 samples/site). As these fully conserved 5’ UTR sequences may have functional importance, we expect that assays targeting this region would broadly detect APPV strains that are diverse in space and time.

scholarly journals The Sua5 Protein Is Essential for Normal Translational Regulation in Yeast

2009 ◽  
Vol 30 (1) ◽  
pp. 354-363 ◽  
Author(s):  
Changyi A. Lin ◽  
Steven R. Ellis ◽  
Heather L. True
Keyword(s):  
Translational Regulation ◽  
Rna Virus ◽  
Critical Role ◽  
Protein Translation ◽  
Nonsense Suppression ◽  
Entry Site ◽  
Stem Loop ◽  
Internal Ribosome Entry ◽  
Reading Frame ◽  
Codon Recognition

ABSTRACT The anticodon stem-loop of tRNAs requires extensive posttranscriptional modifications in order to maintain structure and stabilize the codon-anticodon interaction. These modifications also play a role in accommodating wobble, allowing a limited pool of tRNAs to recognize degenerate codons. Of particular interest is the formation of a threonylcarbamoyl group on adenosine 37 (t6A37) of tRNAs that recognize ANN codons. Located adjacent and 3′ to the anticodon, t6A37 is a conserved modification that is critical for reading frame maintenance. Recently, the highly conserved YrdC/Sua5 family of proteins was shown to be required for the formation of t6A37. Sua5 was originally identified in a screen by virtue of its ability to affect expression from an aberrant upstream AUG codon in the cyc1 transcript. Together, these findings implicate Sua5 in protein translation at the level of codon recognition. Here, we show that Sua5 is critical for normal translation. The loss of SUA5 causes increased leaky scanning through AUG codons, +1 frameshifting, and nonsense suppression. In addition, the loss of SUA5 amplifies the 20S RNA virus found in Saccharomyces cerevisiae, possibly through an internal ribosome entry site-mediated mechanism. This study reveals a critical role for Sua5 and the t6A37 modification in translational fidelity.

scholarly journals Factor-Independent Assembly of Elongation-Competent Ribosomes by an Internal Ribosome Entry Site Located in an RNA Virus That Infects Penaeid Shrimp

2005 ◽  
Vol 79 (2) ◽  
pp. 677-683 ◽  
Author(s):  
Randal C. Cevallos ◽  
Peter Sarnow
Keyword(s):  
Internal Ribosome Entry Site ◽  
Intergenic Region ◽  
Rna Virus ◽  
Penaeid Shrimp ◽  
Initiation Factor ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Ribosomal Subunits ◽  
Initiation Factors ◽  
Functional Conservation

ABSTRACT The Taura syndrome virus (TSV), a member of the Dicistroviridae family of viruses, is a single-stranded positive-sense RNA virus which contains two nonoverlapping reading frames separated by a 230-nucleotide intergenic region. This intergenic region contains an internal ribosome entry site (IRES) which directs the synthesis of the TSV capsid proteins. Unlike other dicistroviruses, the TSV IRES contains an AUG codon that is in frame with the capsid region, suggesting that the IRES initiates translation at this AUG codon by using initiator tRNAmet. We show here that the TSV IRES does not use this or any other AUG codon to initiate translation. Like the IRES in cricket paralysis virus (CrPV), the TSV IRES can assemble 80S ribosomes in the absence of initiation factors and can direct protein synthesis in a reconstituted system that contains only purified ribosomal subunits, eukaryotic elongation factors 1A and 2, and aminoacylated tRNAs. The functional conservation of the CrPV-like IRES elements in viruses that can infect different invertebrate hosts suggests that initiation at non-AUG codons by an initiation factor-independent mechanism may be more prevalent.

scholarly journals Nuclear Protein Sam68 Interacts with the Enterovirus 71 Internal Ribosome Entry Site and Positively Regulates Viral Protein Translation

2015 ◽  
Vol 89 (19) ◽  
pp. 10031-10043 ◽  
Author(s):  
Hua Zhang ◽  
Lei Song ◽  
Haolong Cong ◽  
Po Tien
Keyword(s):  
Life Cycle ◽  
Internal Ribosome Entry Site ◽  
Viral Protein ◽  
Nuclear Protein ◽  
Binding Protein ◽  
Enterovirus 71 ◽  
Protein Translation ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Cellular Factors

ABSTRACTEnterovirus 71 (EV71) recruits various cellular factors to assist in the replication and translation of its genome. Identification of the host factors involved in the EV71 life cycle not only will enable a better understanding of the infection mechanism but also has the potential to be of use in the development of antiviral therapeutics. In this study, we demonstrated that the cellular factor 68-kDa Src-associated protein in mitosis (Sam68) acts as an internal ribosome entry site (IRES)trans-acting factor (ITAF) that binds specifically to the EV71 5′ untranslated region (5′UTR). Interaction sites in both the viral IRES (stem-loops IV and V) and the heterogeneous nuclear ribonucleoprotein K homology (KH) domain of Sam68 protein were further mapped using an electrophoretic mobility shift assay (EMSA) and biotin RNA pulldown assay. More importantly, dual-luciferase (firefly) reporter analysis suggested that overexpression of Sam68 positively regulated IRES-dependent translation of virus proteins. In contrast, both IRES activity and viral protein translation significantly decreased in Sam68 knockdown cells compared with the negative-control cells treated with short hairpin RNA (shRNA). However, downregulation of Sam68 did not have a significant inhibitory effect on the accumulation of the EV71 genome. Moreover, Sam68 was redistributed from the nucleus to the cytoplasm and interacts with cellular factors, such as poly(rC)-binding protein 2 (PCBP2) and poly(A)-binding protein (PABP), during EV71 infection. The cytoplasmic relocalization of Sam68 in EV71-infected cells may be involved in the enhancement of EV71 IRES-mediated translation. Since Sam68 is known to be a RNA-binding protein, these results provide direct evidence that Sam68 is a novel ITAF that interacts with EV71 IRES and positively regulates viral protein translation.IMPORTANCEThe nuclear protein Sam68 is found as an additional new host factor that interacts with the EV71 IRES during infection and could potentially enhance the translation of virus protein. To our knowledge, this is the first report that describes Sam68 actively participating in the life cycle of EV71 at a molecular level. These studies will not only improve our understanding of the replication of EV71 but also have the potential for aiding in developing a therapeutic strategy against EV71 infection.

Apoptosis-Induced Wnt2/β-Catenin Signaling Triggers Astrocytic Dedifferentiation and Facilitates Neurogenesis in Ischemic Cortex

2020 ◽  
Author(s):  
Hong Fan ◽  
Jialei Yang ◽  
Junling Xing ◽  
Baolin Guo ◽  
Wenting Wang ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Caspase 3 ◽  
Protein Translation ◽  
Underlying Mechanism ◽  
Reactive Astrocytes ◽  
Entry Site ◽  
Potential Therapeutic Target ◽  
Internal Ribosome Entry ◽  
Cerebral Ischemic ◽  
Canonical Wnt

Abstract Background: Reactive astrogliosis is a common pathologic change of various neurological disorders and usually takes some properties of neural progenitors. This dedifferentiation response of reactive astrocytes to injury is thought as an endogenous cellular attempt for neuronal regeneration, but the underlying mechanism remains largely unclear. Methods: A focal cerebral ischemic model was adopted to assess the dedifferentiation of reactive astrocytes. Topgal mice (a Wnt signaling reporting mouse line) and Caspase-3-/- mice were used to evaluate the change and roles of Wnt signaling and apoptosis in this process. Virus mediated Wnt2, β-catenin, dnTCF4 and DAP5 manipulation was used to reveal the molecular mechanism of dedifferentiation. Ischemic cortical samples and Wnt2-5UTR sequences of macaca mulatta and human were analyzed to explore if the apoptosis-induced Wnt2 up-regulation was conserved. Results: Focal ischemia induces rapid up-regulation of Wnt2 protein in apoptotic neurons in mice, primates and human, and activation of canonical Wnt signaling in reactive astrocytes. Local delivery of Wnt2 shRNA abolished the dedifferentiation response of astrocytes while over-expressing Wnt2 promoted progenitor marker expression and neurogenesis. Both the activation of Wnt signaling and dedifferentiation of astrocytes was compromised in ischemic caspase-3-/- cortex. Over-expressing stabilized β-catenin not only facilitated neurogenesis but also promoted functional recovery in ischemic caspase-3-/- mice. Apoptotic neurons up-regulated Wnt2 protein via internal ribosome entry site (IRES)-mediated translation. Knocking down death associated protein 5 (DAP5), a key protein in IRES-mediated protein translation, significantly diminished both Wnt activation and astrocyte dedifferentiation. Conclusions: Our data demonstrated a novel apoptosis-initiated Wnt-activating mechanism which triggers the dedifferentiation of reactive astrocytes and facilitates neurogenesis in adult cortex, revealing a “SOS” mechanism for inducing astrocyte dedifferentiation and indicating Wnt2/β-catenin signaling as a potential therapeutic target for ischemic stroke.

scholarly journals EZH2 co-opts gain-of-function p53 mutants to promote cancer growth and metastasis

2018 ◽  
Author(s):  
Yu Zhao ◽  
Liya Ding ◽  
Dejie Wang ◽  
Zhenqing Ye ◽  
Yunqian Pan ◽  
...  
Keyword(s):  
Synthetic Lethality ◽  
P53 Protein ◽  
Protein Translation ◽  
Cancer Growth ◽  
Common Mechanism ◽  
Entry Site ◽  
Gain Of Function ◽  
Independent Manner ◽  
Internal Ribosome Entry ◽  
P53 Mrna

AbstractWith the unfolding of more and more cancer-driven gain-of-function (GOF) mutants of p53, it is important to define a common mechanism to systematically target different mutants rather than develop strategies tailored to inhibit each mutant individually. Here, using RNA immunoprecipitation sequencing (RIP-seq) we identified EZH2 as a p53 mRNA-binding protein. EZH2 bound to the internal ribosome entry site (IRES) in the 5’ untranslated region (5’UTR) of p53 mRNA and enhanced p53 protein translation in a methyltransferase-independent manner. EZH2 augmented p53 GOF mutant-mediated cancer growth and metastasis by increasing p53 GOF mutant protein level. EZH2 overexpression associated with the worse outcome only in patients with p53-mutated cancer. Depletion of EZH2 by antisense oligonucleotides inhibited p53 GOF mutant-mediated cancer growth. Our findings reveal a non-methyltransferase function of EZH2 that controls protein translation of p53 GOF mutants, inhibition of which causes synthetic lethality in cancer cells expressing p53 GOF mutants.

scholarly journals SARS-CoV-2 mutations altering regulatory properties: deciphering host’s and virus’s perspectives

2020 ◽  
Author(s):  
Abul Bashar Mir Md. Khademul Islam ◽  
Md. Abdullah-Al-Kamran Khan
Keyword(s):  
Enrichment Analysis ◽  
Functional Enrichment Analysis ◽  
Functional Enrichment ◽  
Functional Importance ◽  
Entry Site ◽  
Protein Coding ◽  
Internal Ribosome Entry ◽  
The World ◽  
Regulatory Properties ◽  
Different Parts

AbstractSince the first recorded case of the SARS-CoV-2, it has acquired several mutations in its genome while spreading throughout the globe. However, apart from some changes in protein coding, functional importance of these mutations in disease pathophysiology are still largely unknown. In this study, we investigated the significance of these mutations both from the host’s and virus’s perspective by analyzing the host miRNA binding and virus’s internal ribosome entry site (IRES), respectively. Strikingly, we observed that due to the acquired mutations, host miRNAs bind differently compared to the reference; where few of the miRNAs lost and few gained the binding affinity for targeting the viral genome. Moreover, functional enrichment analysis suggests that targets of both of these gained and lost miRNAs might be involved in various host immune signaling pathways. Also, we sought to shed some insights on the impacts of mutations on the IRES structure of SARS-CoV-2. Remarkably, we detected that three particular mutations in the IRES can disrupt its secondary structure which can further make the virus less functional. These results could be valuable in exploring the functional importance of the mutations of SARS-CoV-2 and could provide novel insights into the differences observed different parts of the world.

scholarly journals DDX21, a Host Restriction Factor of FMDV IRES-Dependent Translation and Replication

Viruses ◽  
2021 ◽  
Vol 13 (9) ◽  
pp. 1765
Author(s):  
Sahibzada Waheed Abdullah ◽  
Jin’en Wu ◽  
Yun Zhang ◽  
Manyuan Bai ◽  
Junyong Guan ◽  
...  
Keyword(s):  
Ribosome Biogenesis ◽  
Foot And Mouth Disease ◽  
Protein Translation ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Host Restriction ◽  
Protein Protein Interaction ◽  
Host Restriction Factor ◽  
Pulldown Assay ◽  
Mouth Disease Virus

In cells, the contributions of DEAD-box helicases (DDXs), without which cellular life is impossible, are of utmost importance. The extremely diverse roles of the nucleolar helicase DDX21, ranging from fundamental cellular processes such as cell growth, ribosome biogenesis, protein translation, protein–protein interaction, mediating and sensing transcription, and gene regulation to viral manipulation, drew our attention. We designed this project to study virus–host interactions and viral pathogenesis. A pulldown assay was used to investigate the association between foot-and-mouth disease virus (FMDV) and DDX21. Further insight into the DDX21–FMDV interaction was obtained through dual-luciferase, knockdown, overexpression, qPCR, and confocal microscopy assays. Our results highlight the antagonistic feature of DDX21 against FMDV, as it progressively inhibited FMDV internal ribosome entry site (IRES) -dependent translation through association with FMDV IRES domains 2, 3, and 4. To subvert this host helicase antagonism, FMDV degraded DDX21 through its non-structural proteins 2B, 2C, and 3C protease (3Cpro). Our results suggest that DDX21 is degraded during 2B and 2C overexpression and FMDV infection through the caspase pathway; however, DDX21 is degraded through the lysosomal pathway during 3Cpro overexpression. Further investigation showed that DDX21 enhanced interferon-beta and interleukin-8 production to restrict viral replication. Together, our results demonstrate that DDX21 is a novel FMDV IRES trans-acting factor, which negatively regulates FMDV IRES-dependent translation and replication.

scholarly journals Complete genome sequence analysis of Seneca Valley virus-001, a novel oncolytic picornavirus

2008 ◽  
Vol 89 (5) ◽  
pp. 1265-1275 ◽  
Author(s):  
Laura M. Hales ◽  
Nick J. Knowles ◽  
P. Seshidar Reddy ◽  
Ling Xu ◽  
Carl Hay ◽  
...  
Keyword(s):  
Genome Sequence ◽  
Complete Genome Sequence ◽  
Complete Genome ◽  
Rna Virus ◽  
Encephalomyocarditis Virus ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Reading Frame ◽  
Encephalomyelitis Virus ◽  
Seneca Valley Virus

The complete genome sequence of Seneca Valley virus-001 (SVV-001), a small RNA virus, was determined and was shown to have typical picornavirus features. The 7280 nt long genome was predicted to contain a 5′ untranslated region (UTR) of 666 nt, followed by a single long open reading frame consisting of 6543 nt, which encodes a 2181 aa polyprotein. This polyprotein could potentially be cleaved into 12 polypeptides in the standard picornavirus L-4-3-4 layout. A 3′ UTR of 71 nt was followed by a poly(A) tail of unknown length. Comparisons with other picornaviruses showed that the P1, 2C, 3C and 3D polypeptides of SVV-001 were related most closely to those of the cardioviruses, although they were not related as closely to those of encephalomyocarditis virus and Theiler's murine encephalomyelitis virus as the latter were to each other. Most other regions of the polyprotein differed considerably from those of all other known picornaviruses. SVV-001 contains elements of an internal ribosome entry site reminiscent of that found in hepatitis C virus and a number of genetically diverse picornaviruses. SVV-001 is a novel picornavirus and it is proposed that it be classified as the prototype species in a novel genus named ‘Senecavirus’.

scholarly journals A Highly Divergent Picornavirus in a Marine Mammal

2007 ◽  
Vol 82 (1) ◽  
pp. 311-320 ◽  
Author(s):  
A. Kapoor ◽  
J. Victoria ◽  
P. Simmonds ◽  
C. Wang ◽  
R. W. Shafer ◽  
...  
Keyword(s):  
Marine Mammal ◽  
Rna Virus ◽  
Phylogenetic Position ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Seal Population ◽  
Ljungan Virus ◽  
High Prevalence ◽  
Protein Sequence Identity ◽  
High Level

ABSTRACT Nucleic acids from an unidentified virus from ringed seals (Phoca hispida) were amplified using sequence-independent PCR, subcloned, and then sequenced. The full genome of a novel RNA virus was derived, identifying the first sequence-confirmed picornavirus in a marine mammal. The phylogenetic position of the tentatively named seal picornavirus 1 (SePV-1) as an outlier to the grouping of parechoviruses was found consistently in alignable regions of the genome. A mean protein sequence identity of only 19.3 to 30.0% was found between the 3D polymerase gene sequence of SePV-1 and those of other picornaviruses. The predicted secondary structure of the short 506-base 5′-untranslated region showed some attributes of a type IVB internal ribosome entry site, and the polyprotein lacked an apparent L peptide, both properties associated with the Parechovirus genus. The presence of two SePV-1 2A genes and of the canonical sequence required for cotranslational cleavage resembled the genetic organization of Ljungan virus. Minor genetic variants were detected in culture supernatants derived from 8 of 108 (7.4%) seals collected in 2000 to 2002, indicating a high prevalence of SePV-1 in this hunted seal population. The high level of genetic divergence of SePV-1 compared to other picornaviruses and its mix of characteristics relative to its closest relatives support the provisional classification of SePV-1 as the prototype for a new genus in the family Picornaviridae.

scholarly journals Ectropis obliqua picorna-like virus IRES-driven internal initiation of translation in cell systems derived from different origins

2007 ◽  
Vol 88 (10) ◽  
pp. 2834-2838 ◽  
Author(s):  
Jie Lu ◽  
Yuanyang Hu ◽  
Liu Hu ◽  
Shan Zong ◽  
Dawei Cai ◽  
...  
Keyword(s):  
Translation Initiation ◽  
Rna Virus ◽  
Translation System ◽  
Entry Site ◽  
Host Preferences ◽  
Internal Ribosome Entry ◽  
Initiation Of Translation ◽  
Internal Initiation ◽  
Internal Translation ◽  
Different Origins

Ectropis obliqua picorna-like virus (EoPV) is an insect RNA virus that causes a lethal granulosis infection of larvae of the tea looper (Ectropis obliqua). An internal ribosome entry site (IRES) mediates translation initiation of EoPV RNA. Here, bicistronic constructs were used to examine the 5′ untranslated region (UTR) of EoPV for IRES activity. The capacities of the EoPV 5′ UTR IRES and another insect virus IRES, the cricket paralysis virus intergenic region IRES, to mediate internal translation initiation in a variety of translation systems were also compared. The results demonstrated that the EoPV IRES functioned efficiently not only in mammalian cell-derived systems, but also in an insect cell-derived translation system. However, it functioned inefficiently in a plant cell-derived translation system. This study reveals the host preferences of the EoPV IRES and important differences in IRES function between the EoPV IRES and other characterized picorna-like insect viral IRESs.

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